| igDirect {CalciOMatic} | R Documentation |
The function igDirect provides an initial guess for the
experiment-specific parameters of fluorescence transients obtained
with a ratiometric dye (the background fluorescence log_S_B_340
and log_S_B_380, as well as the scaling coefficient
log_phi)
igDirect(adu_B_340, adu_340, adu_B_380, adu_380,
ig_ratio, t, tOn = 1, subset = 1:length(t),
R_min = 0.136, R_max = 2.701, K_eff = 3.637, K_d = 0.583,
B_T = 100, T_340 = 0.015, T_380 = 0.006, P = 400, P_B = 400)
adu_B_340 |
the background fluorescence at 340 nm |
adu_340 |
the fluorescence transient at 340 nm |
adu_B_380 |
the background fluorescence at 380 nm |
adu_380 |
the fluorescence transient at 380 nm |
ig_ratio |
the initial guess list for the parameters of the
Ca^2+ transient, returned by the igRatio
function |
t |
a vector of time values at which the fluorescence values were obtained (in s) |
tOn |
the time of the fluorescence jump (in s) |
subset |
a vector of time indices to consider (generally the whole fluorescence signals) |
R_min |
the minimum fluorescence ratio between the measurements at 340 and 380 nm. This parameter is obtained from calibration experiments |
R_max |
the maximum fluorescence ratio between the measurements at 340 and 380 nm. This parameter is obtained from calibration experiments |
K_eff |
the effective dissociation constant of the dye in the cell (in muM). This parameter is obtained from calibration experiments |
K_d |
the dissociation constant of the dye in the cell (in muM). This parameter is obtained from calibration experiments |
B_T |
the total concentration of the dye in the cell (in muM) |
T_340 |
the exposure time at 340 nm (in s) |
T_380 |
the exposure time at 380 nm (in s) |
P |
the number of pixels of the Region Of Interest (ROI) |
P_B |
the number of pixels of the Background Region |
The intial guesses for log_S_B_340 and log_S_B_380 are
obtained by averaging the adu_B_340 and adu_B_380
signals, divided by T_340*P_B and T_380*P_B
respectively, and by taking their logarithm.
The initial guess for log_phi is obtained by applying a linear
(zero-intercept) regression between the following signals, and by
taking the logarithm of the slope:
c(adu_340/T_340/P - S_B_340, adu_380/T_380/P - S_B_380)
B_T / (K_d+Ca_ratio) * c(R_min*K_eff+R_max*Ca_ratio,K_eff+Ca_ratio)
In these formulas, Ca_ratio refers to the
calcium concentration transient estimated with the initial guess of
the parameters listed in the ig_ratio argument
A named list of class "initial_guess", containing initial
guesses (IG) for the logarithms of the three experiment-specific
parameters: The background fluorescences at 340 and 380 nm
(log_S_B_340 and log_S_B_380 respectively) and the
amplitude coefficient log_phi
Sebastien Joucla sebastien.joucla@parisdescartes.fr
see the fluo documentation for details
about the data generation model
## Parameters of the monoexponential calcium transient
tOn <- 1
Time <- seq(0,12,length.out=160)
Ca0 <- 0.10
dCa <- 0.25
tau <- 1.5
## Create a monoexponential calcium decay
Ca <- caMonoExp(t = Time, tOn = tOn,
Ca0 = Ca0, dCa = dCa, tau = tau)
## Calibrated parameters
R_min <- list(value=0.136, mean=0.136, se=0.00363, USE_se=TRUE)
R_max <- list(value=2.701, mean=2.701, se=0.151, USE_se=TRUE)
K_eff <- list(value=3.637, mean=3.637, se=0.729, USE_se=TRUE)
K_d <- list(value=0.583, mean=0.583, se=0.123, USE_se=TRUE)
## Experiment-specific parameters
nb_B <- 1
B_T <- 100.0
T_340 <- 0.015
T_380 <- 0.006
P <- 200
P_B <- 200
phi <- 2
S_B_340 <- 30
S_B_380 <- 80
## Define Background and Transient fluorescence
## signals at 340 and 380 nm
adu_B_340 <- fluo(Ca=rep(0,nb_B),
R_min=R_min$value, R_max=R_max$value,
K_eff=K_eff$value, K_d=K_d$value,
B_T=0, phi=phi, S_B=S_B_340,
T_stim=T_340, P=P, P_B=P_B)
adu_340 <- fluo(Ca=Ca,
R_min=R_min$value, R_max=R_max$value,
K_eff=K_eff$value, K_d=K_d$value,
B_T=B_T, phi=phi, S_B=S_B_340,
T_stim=T_340, P=P, P_B=P_B)
adu_B_380 <- fluo(Ca=rep(0,nb_B),
R_min=1, R_max=1,
K_eff=K_eff$value, K_d=K_d$value,
B_T=0, phi=phi, S_B=S_B_380,
T_stim=T_380, P=P, P_B=P_B)
adu_380 <- fluo(Ca=Ca,
R_min=1, R_max=1,
K_eff=K_eff$value, K_d=K_d$value,
B_T=B_T, phi=phi, S_B=S_B_380,
T_stim=T_380, P=P, P_B=P_B)
## Add Poissonian noise to these signals
adu_B_340 <- rpois(length(adu_B_340), adu_B_340)
adu_340 <- rpois(length(adu_340), adu_340)
adu_B_380 <- rpois(length(adu_B_380), adu_B_380)
adu_380 <- rpois(length(adu_380), adu_380)
## Extract the noisy calcium transient
## (from the ratiometric transformation)
Ca_noisy <- caFromRatio(adu_B_340, adu_340,
adu_B_380, adu_380,
T_340, T_380,
P, P_B,
R_min, R_max, K_eff,
Plot = FALSE)
## Perform a ratiometric fit to determine
## the calcium dynamics parameters
ratio_fit <- ratioFitFromCa(Ca_noisy, t=Time, tOn, type="mono")
## List the fitted parameters and create
## the corresponding calcium transient
ig_mono <- as.list(ratio_fit$par)
names(ig_mono) <- c("log_Ca0","log_dCa","log_tau")
class(ig_mono) <- "initial_guess"
## Perform an Initial Guess for the Experiment-Specific Parameters
ig_direct <- igDirect(adu_B_340 = adu_B_340,
adu_340 = adu_340,
adu_B_380 = adu_B_380,
adu_380 = adu_380,
ig_ratio = ig_mono,
t = Time, tOn = tOn, subset = 1:length(Time),
R_min = R_min$value, R_max = R_max$value,
K_eff = K_eff$value, K_d = K_d$value,
B_T = 100, T_340 = T_340, T_380 = T_380, P = P, P_B = P_B)
## Compare the initial guess with the known values of the parameters
print(exp(as.vector(unlist(ig_direct))))
print(c(Ca0, dCa, tau, phi, S_B_340, S_B_380))